The present invention relates to a support for the determination of enzyme activities and to a method for such determination employing said support.
It is known that paper discs may be used to perform chemical tests, such as strip tests for albumin, glucose and or disc tests for antibiotics, ONPG and the like wherein the reagent is simply placed on a sheet of paper. These are approximate methods, and do not serve to provide quantitative results.
Traditional methods of precision analysis call for complicated and costly apparatus and are time-consuming.
For the conventional determination of enzyme activity, it has been necessary to prepare a purified biological extract and an aqueous buffer with a pH corresponding to the pH of the activity of the enzyme, for each activity. This means that as many buffers must be prepared as there are optimum pH-values of activity, when several enzymes acting at different pH-values are to be determined.
Moreover, if the substrate is to be dissolved in the buffer, the use of substrates insoluble in water is excluded. If the substrate is not directly chromogenic, a coloring agent must be prepared and the so-formed colored product of the determination must itself be soluble in aqueous media. This excludes all substrates that yield water-insoluble reaction products. In addition, for each activity tested, the substrate, the biological extract in the buffer and other reagents such as the coloring agent must be mixed in proper proportions. Finally, since these reactions are determined spectrophotometrically, a photometer must be employed.
Besides the need for comparatively costly apparatus, such a procedure is very time-consuming in practice, since as a rule no result can be obtained until after several days of experiments carried out by a qualified technician. It also has the disadvantage of requiring a comparatively large quantity of the biological extracts to be tested.
The present invention on the other hand provides a novel apparatus and method to perform the determination of enzyme activities quickly and simply. It is not the purpose of the present invention to attain the precision of spectrophotometric determinations for each enzyme, but it is intended essentially for detection of enzyme activities in an unpurified complex extract. Neither is the invention intended to take the place of electrophoretic separating methods. The present invention rather serves as an excellent guide for the latter by previously giving the spectrum of enzyme activities of the sample to be tested in its crude state.